Studies on a cellular extract (D98) have continued. We have found that the D98 extract can inhibit EBV DNA synthesis by approximately 1/3 in superinfected cells as compared to the control. The inhibitory factor is sensitive to trypsin but not DNase or RNase. In addition, the inhibitory portion of the extract is nondialyzable and has a molecular weight of greater than 10,000. Herpes simplex type 1 and type 2 are not inhibited by the D98 extract and no evidence associating this factor(s) with an interferon like protein has been found. We are studying the molecular events associated with the replication of HR-1 EBV in EBV genome positive and genome negative lymphoblastoid cell lines. Both cell types can be superinfected, but thus far, only transforming virus has been obtained from superinfected Raji cells. EBV DNA synthesis was observed in virus genome negative Ramos cells after superinfection, thus endogenous EBV DNA is not required for virus replication. The Eco R1 and Sma I restriction enzyme patterns of EBV DNA are similar in superinfected genome positive and negative nonproducer cell lines. Studies on early and late transcription of the EBV genome have been started using control and phosphonoacetic acid treated infected cells and potassium iodide (KI) density gradients. Evidence has been found supporting temporal regulation of EBV transcription during lytic infection. EBV DNA synthesized early after infection showed a higher S value than linear 55S EBV DNA. At late stage, homogeneous unit length DNA was detected.